1. Field of the Invention
The present invention relates to the field of molecular biology and genetic engineering, and polynucleic acid molecules useful for gene expression in prokaryotes. Specifically, the present invention discloses chimeric polynucleic acid molecules comprising promoter activity in bacterial cells. The invention further discloses DNA constructs and bacterial cells comprising the polynucleic acid molecules, and methods of producing and using the same.
2. Description of Related Art
One of the goals of genetic engineering is to produce organisms with desirable characteristics or traits. The proper expression of a desirable transgene in a transgenic organism is one way to achieve this goal. Elements having gene regulatory activity, i.e. regulatory elements such as promoters, leaders, and transcription termination regions, are non-coding polynucleotide molecules that play an integral part in the overall expression of genes in living cells. Regulatory elements that function in prokaryotes are therefore useful for modifying their phenotypes through the methods of genetic engineering.
While previous work has provided a number of regulatory elements useful to affect gene expression in transgenic bacteria, there is still a great need for novel regulatory elements with beneficial expression characteristics. Many previously identified regulatory elements fail to provide the patterns or levels of expression required to fully realize the benefits of expression of selected genes in transgenic bacteria. One example of this is the need for regulatory elements capable of driving strong gene expression in different types of bacteria.
A promoter is a key element for directing gene expression in a cell. The transcription machinery is assembled and transcription is initiated from the promoter DNA molecule. Transcription factors influence the strength of a transcript from a promoter molecule. Accordingly, regions within the promoter molecule function to enhance or repress transcription.
The genetic enhancement of bacteria provides significant benefits to society. For example, bacteria may be enhanced with a transgene to provide desirable biosynthetic, commercial, chemical, insecticidal, industrial, nutritional, or pharmaceutical properties. Despite the availability of many molecular tools, however, the genetic modification of bacteria is often constrained by an insufficient expression of the engineered transgene.
High level gene expression requires a strong 5′ regulatory sequence (promoter) and may also be affected by sequences found 3′ to a coding sequence. Currently only a limited number of strong promoters are available from prokaryotes. Thus, there is a need for additional promoters that are useful for expressing genes, especially single copy genes in a single or low copy vector.